title: Comparison of the membrane proteome of virulent Mycobacterium tuberculosis and the attenuated Mycobacterium bovis BCG vaccine strain by label-free quantitative proteomics.
abstract: The Mycobacterium tuberculosis membrane is rich in antigens that are potential targets for diagnostics and the development of new vaccines. To better understand the mechanisms underlying MTB virulence and identify new targets for therapeutic intervention, we investigated the differential composition of membrane proteomes between virulent M. tuberculosis H37Rv (MTB) and the Mycobacterium bovis BCG vaccine strain. To compare the membrane proteomes, we used LC-MS/MS analysis in combination with label-free quantitative proteomics, utilizing the area under the curve of the extracted ion chromatograms of peptides obtained from m/z and retention time alignment of MS1 features. With this approach, we obtained relative abundance ratios for 2203 identified membrane-associated proteins in high confidence. Of these proteins, 294 showed statistically significant differences of at least two fold in relative abundance between MTB and BCG membrane fractions. Our comparative analysis detected several proteins associated with known genomic regions of difference between MTB and BCG as being absent, which validated the accuracy of our approach. In further support of our label-free quantitative data, we verified select protein differences by immunoblotting. To our knowledge, we have generated the first comprehensive and high-coverage profile of comparative membrane proteome changes between virulent MTB and its attenuated relative BCG, which helps elucidate the proteomic basis of the intrinsic virulence of the MTB pathogen.
authors.affiliation: Department of Biochemistry and Biophysics, University of North Carolina at Chapel Hill , 120 Mason Farm Road, Campus Box 7260 3rd Floor, Genetic Medicine Building, Chapel Hill, North Carolina 27599, United States.
authors.forename: Harsha P
authors.initials: HP
authors.lastname: Gunawardena
authors.forename: Meghan E
authors.initials: ME
authors.lastname: Feltcher
authors.forename: John A
authors.initials: JA
authors.lastname: Wrobel
authors.forename: Sheng
authors.initials: S
authors.lastname: Gu
authors.forename: Miriam
authors.initials: M
authors.lastname: Braunstein
authors.forename: Xian
authors.initials: X
authors.lastname: Chen
mesh_terms: D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification; D002462:Cell Membrane / Q000737:chemistry; D002853:Chromatography, Liquid; D015964:Gene Expression Regulation, Bacterial; D056426:Genetic Loci; D015151:Immunoblotting; D008565:Membrane Proteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification; D058977:Molecular Sequence Annotation; D009163:Mycobacterium bovis / Q000737:chemistry* / Q000235:genetics; D009169:Mycobacterium tuberculosis / Q000737:chemistry* / Q000235:genetics / Q000472:pathogenicity*; D010455:Peptides; D059748:Proteolysis; D020543:Proteome / Q000737:chemistry*; D040901:Proteomics / Q000379:methods; D053719:Tandem Mass Spectrometry; D014357:Trypsin / Q000737:chemistry; D014774:Virulence
chemical_list: D001426:Bacterial Proteins; D008565:Membrane Proteins; D010455:Peptides; D020543:Proteome; D014357:Trypsin